In this project we have been paricularly interested in thephysiologic function of the large amount of polyamines normally present in cells. For this purpose in an ongoing project we have performed microarray studies in yeast polyamine requiring mutants to find out specific effects of spermidine or spermine addition or depletion on global gene expression (with the help of the NIDDK Microarray Facility). The analysis of the data revealed that spermudine has more effect on transcription of yeast genes than spermine. In a study with a deleted spe3 S. cerevisiae auxotroph we found that spermidine addition resulted in more than a4-fold induction of 73 genes including some of the transcription factors (MET28, GAT1). Functional categorization of the genes showed induction of transport related genes (16%), and amino acid metabolism, in particular methionine biosynthesis (19%). Spermidine also repressed 28 genes more than 4-fold. In contrast, only few genes (14) were significantly responsive to spermine. Real time PCR analysis o fa few represetnative genes showed a similar trend to that found in the microarray experiments.[unreadable] [unreadable] Further characterization of specific pathways and their metabolites affected by spermidine addition are under investigation using other mutants deleted in polyamine biosynthesis with a particular interest in studing why yeast cells normally contain much higher concentrations of polyamines than needed for growth.. In particular we have recently carried out such studies on the early and late responses of spermidine additions using a yeast double mutants (deleted spe1-deleted spe 2) We have detected several interesting targets of spermidine, which include several transcripts of heat shock proteins, zinc and copper transporters, zinc finger transcription factors, TyA (Gag protein) and TyB (Gag-Pol) proteins.[unreadable] [unreadable] We are also continuing our studies on the role of spermidine in protecting polyamine auxotrophs from oxygen toxicity, with particular interest in the level of spermidine required.